Biofilm-related infections caused by Staphylococcus epidermidis and Staphylococcus aureus pose significant medical challenges. They are the leading cause of hospital-acquired infections, affecting up to 400,000 patients/year in the United States. In order to address this unmet medical need, we are developing small molecules that specifically inhibit staphylococcal biofilm formation. These novel drugs will be used as coatings for indwelling medical devices, such as central venous catheters, to prevent staphylococcal biofilm infections. In a previous Phase II SBIR grant, we identified a class of small molecules, known as the rhodanines, that are potent inhibitors of in vitro staphylococcal biofilm formation. In Phase I of this SBIR, we will optimize the rhodanine biofilm inhibitors for the generation of a Lead compound. Repeated rounds of synthesis and evaluation in in vitro and in vivo assays will be used to explore the structure activity relationships of this chemical class and to improve potency and selectivity. Compounds that meet the specified criteria for anti-biofilm potency in vitro (MBIC = 0.1 [unreadable]M), specificity (MIC/MBIC = 500), and selectivity (CC50/MBIC = 50) will be advanced to mechanism of action (MOA) studies and in vivo assays for toxicity and efficacy. Compounds with verified MOA, in vivo efficacy, and acceptable in vivo toxicity (MTD<1mg/kg) will be designated as Lead compounds. In Phase II, Lead compounds will be optimized using rational drug design. The Specific Aims for this Phase I proposal are as follows. Aim 1. Design and synthesize rhodanine analogs with improved anti-biofilm activity. Aim 2. Optimize in vitro anti-biofilm activity, spectrum, and selectivity of novel inhibitors of biofilm formation. Aim 3. Determine in vitro anti- biofilm activity of catheter material impregnated with anti-biofilm compounds. Aim 4. Determine mechanism of action (MOA) of optimized compounds. Aim 5. Determine acute toxicity and efficacy of lead compounds in an in vivo model for biofilm infection. [unreadable] [unreadable] The goal of this proposal is to develop drugs that will be used as coatings on medical devices that prevent staphylococcal biofilm formation. This is important because Staphylococcus epidermidis and Staphylococcus aureus are responsible for the vast majority of biofilm-related infections associated with the use of indwelling medical devices. Staphylococcal pathogens are able to colonize the surfaces of medical devices in the human body and form biofilms, which represents a unique mode of surface-attached growth. Bacteria growing in biofilms are highly resistant to antibiotics, biocides, and attack by the body's immune system. As a result, biofilm infections are difficult to eradicate and lead to recurrent bloodstream infections. The innovation of this proposal is to develop drugs that specifically target the biofilm mode of growth to prevent biofilm-related infections. [unreadable] [unreadable] [unreadable]